1) Dr. Aravind and his group performed research to identify novel nucleic acid modification enzymes. Modified bases in nucleic acids present a layer of information that directs biological function over and beyond the coding capacity of the conventional bases. Recently, members of the 2-oxoglutarate- and iron(II)-dependent dioxygenase super-family, which modify diverse substrates from small molecules to biopolymers, were predicted and subsequently confirmed to catalyze oxidative modification of bases in nucleic acids. Of these, two distinct families, namely the AlkB and the kinetoplastid base J binding proteins (JBP) catalyze in situ hydroxylation of bases in nucleic acids. Using sensitive computational analysis of sequences, structures and contextual information from genomic structure and protein domain architectures, they reported five distinct families of 2-oxoglutarate- and iron(II)-dependent dioxygenase that they predict to be involved in nucleic acid modifications. Among the DNA-modifying families, they showed that the dioxygenase domains of the kinetoplastid base J-binding proteins belong to a larger family that includes the Tet proteins, prototyped by the human oncogene Tet1, and proteins from basidiomycete fungi, chlorophyte algae, heterolobosean amoeboflagellates and bacteriophages. They presented evidence that some of these proteins are likely to be involved in oxidative modification of the 5-methyl group of cytosine leading to the formation of 5-hydroxymethylcytosine. The Tet/JBP homologs from basidiomycete fungi such as Laccaria and Coprinopsis show large lineage-specific expansions and a tight linkage with genes encoding a novel and distinct family of predicted transposases, and a member of the Maelstrom-like HMG family. They proposed that these fungal members are part of a mobile transposon. This is the first report of a eukaryotic transposable element that encodes its own DNA-modification enzyme with a potential regulatory role. Through a wider analysis of other poorly characterized DNA-modifying enzymes they also showed that the phage Mu Mom-like proteins, which catalyze the N6-carbamoylmethylation of adenines, are also linked to diverse families of bacterial transposases, suggesting that DNA modification by transposable elements might have a more general presence than previously appreciated. Among the other families of 2-oxoglutarate- and iron(II)-dependent dioxygenases identified in this study, one which is found in algae, is predicted to mainly comprise of RNA-modifying enzymes and shows a striking diversity in protein domain architectures suggesting the presence of RNA modifications with possibly unique adaptive roles. The results presented here are likely to provide the means for future investigation of unexpected epigenetic modifications, such as hydroxymethyl cytosine, that could profoundly impact our understanding of gene regulation and processes such as DNA demethylation. 2)DNA cytosine methylation is crucial for retrotransposon silencing and mammalian development. In a computational search for enzymes that could modify 5-methylcytosine (5mC), Dr. Aravind identified TET proteins as mammalian homologs of the trypanosome proteins JBP1 and JBP2, which have been proposed to oxidize the 5-methyl group of thymine. In conjunction with his collaborator Dr. Anjana Rao, Immune Disease Institute, Harvard Medical School it was shown that TET1, a fusion partner of the MLL gene in acute myeloid leukemia, is a 2-oxoglutarate (2OG)- and Fe(II)-dependent enzyme that catalyzes conversion of 5mC to 5-hydroxymethylcytosine (hmC) in cultured cells and in vitro. hmC is present in the genome of mouse embryonic stem cells, and hmC levels decrease upon RNA interference-mediated depletion of TET1. Thus, TET proteins have potential roles in epigenetic regulation through modification of 5mC to hmC. 3)Dr. Aravind initiated a project on chromatin proteins that lead to the discovery of a novel histone chaperone that function as part of the Hir-Asf1 histone chaperone complex. Using protein sequence profile analyses methods he investigated the evolution of Hir-Asf1 complex proteins and showed that they have a much wider phyletic pattern than was previously known. He established the animal histone-deacetylase-complex-interacting proteins, CAIN/CABIN, to be orthologs of Hir3p. They contain a conserved core of around 30 TPR-like bi-helical repeats that are likely to form a super-helical scaffold. He also identified a conserved domain, the HUN domain, in all Hpc2p homologs, including animal ubinuclein/yemanuclein and the recently discovered vertebrate cell-cycle regulator FLJ25778. The HUN domain has a characteristic pattern of conserved acidic residues based on which he predicted that it is a previously unrecognized histone-tail-binding chaperone. By analyzing various high-throughput data sets, such as RNAi knock-downs, genetic and protein interaction maps and cell-cycle-specific gene expression data, we present evidence that Hpc2p homologs might be deployed in specific processes of chromatin dynamics relating to cell-cycle progression in vertebrates and schizogony in Plasmodium.